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Biomedical subjects

C M Preston

Publications and source records attributed to C M Preston.

At least 19 recordsLinked to original sources
PubMed2005

Towards an understanding of the molecular basis of herpes simplex virus latency.

The survival strategy of herpes simplex virus centres on the establishment of latency in sensory neurons innervating the site of primary infection followed by periodic reactivation to facilitate transmission. This is a highly evolved and efficient survival mechanism, which despite being the subject of intense research, has proven remarkably difficult to dissect at a molecular level. This review will focus on data, emerging from both in vitro and in vivo model systems, which provide a framework for a mechanistic understanding of latency and the existence and possible significance of non-uniform latent states.

Animals↗
PubMed2004

Phylogenetic screening of ribosomal RNA gene-containing clones in Bacterial Artificial Chromosome (BAC) libraries from different depths in Monterey Bay.

Marine picoplankton are central mediators of many oceanic biogeochemical processes, but much of their biology and ecology remains ill defined. One approach to better defining these environmentally significant microbes involves the acquisition of genomic data that can provide information about genome content, metabolic capabilities, and population variability in picoplankton assemblages. Previously, we constructed and phylogenetically screened a Bacterial Artificial Chromosome (BAC) library from surface water picoplankton of Monterey Bay. To further describe niche partitioning, metabolic variability, and population structure in coastal picoplankton populations, we constructed and compared several picoplankton BAC libraries recovered from different depths in Monterey Bay. To facilitate library screening, a rapid technique was developed (ITS-LH-PCR) to identify and quantify ribosomal RNA (rRNA) gene-containing BAC clones in BAC libraries. The approach exploited natural length variations in the internal transcribed spacer (ITS) located between SSU and LSU rRNA genes, as well as the presence and location of tRNA-alanine coding genes within the ITS. The correspondence between ITS-LH-PCR fragment sizes and 16S rRNA gene phylogenies facilitated rapid identification of rRNA genes in BAC clones without requiring direct DNA sequencing. Using this approach, 35 phylogenetic groups (previously identified by cultivation or PCR-based rRNA gene surveys) were detected and quantified among the BAC clones. Since the probability of recovering chimeric rRNA gene sequences in large insert BAC clones was low, we used these sequences to identify potentially chimeric sequences from previous PCR amplified clones deposited in public databases. Full-length SSU rRNA gene sequences from picoplankton BAC libraries, cultivated bacterioplankton, and nonchimeric RNA genes were then used to refine phylogenetic analyses of planktonic marine gamma Proteobacteria, Roseobacter, and Rhodospirillales species.

Bacteria↗
PubMed2003

Linking chemical reactivity and protein precipitation to structural characteristics of foliar tannins.

Tannins influence ecosystem function by affecting decomposition rates, nutrient cycling, and herbivory. To determine the role of tannins in ecological processes, it is important to quantify their abundance and understand how structural properties affect reactivity. In this study, purified tannins from the foliage of nine species growing in the pygmy forest of the northern California coast were examined for chemical reactivity, protein precipitation capacity (PPC), and structural characteristics (13C NMR). Reactivity of purified tannins varied among species 1.5-fold for the Folin total phenol assay, and 7-fold and 3-fold, respectively, for the acid butanol and vanillin condensed tannin assays. There was about a 5-fold difference in PPC. Variation in chemical reactivity and PPC can be largely explained by differences in structural characteristics of the tannins determined by 13C NMR. In particular, the condensed versus hydrolyzable tannin content, as well as the hydroxylation pattern of the B-ring and stereochemistry at the C-2-C-3 position appear to influence reactivity. Due to the large differences in chemical reactivity among species, it is necessary to use a well-characterized purified tannin from the species of interest to convert assay values to concentrations. Our results suggest that structural characteristics of tannins play an important role in regulating their reactivity in ecological processes.

Animals↗
PubMed2002

Effects of wheat variety and specific weight on dietary apparent metabolisable energy concentration and performance of broiler chicks.

1. This study used 4 wheat cultivars (Brigadier, Chaucer, Consort, Reaper) from three locations (Crossnacreevy, C; Downpatrick, D; Limavady, L), which had given rise to differences in wheat specific weight (SW), to examine the relationships between apparent metabolisable energy (AME) concentration, broiler performance and wheat SW. 2. The diets contained (g/kg): wheat 744, casein 142, blended vegetable fat 50, dicalcium phosphate 22, potassium bicarbonate 10.8, sodium bicarbonate 7.5, arginine 5, methionine 2, binder 8, trace minerals/vitamins 7.2, titanium dioxide 1.5. The diets were heat-treated (80 degrees C for 2 min) prior to pelleting (3 mm die). 3. SW ranged from 63 to 77 kg/hectolitre (hl), averaging 66, 69 and 76 kg/hl at D, C and L, respectively. In vitro viscosity of the wheat samples ranged from 5.2 to 17.5 cps and thousand grain weight (TGW) from 33.4 to 47.3 g. Mean TGW was similar at C and D (38.7, 37.0 g) but higher at L (43.1 g). In vitro viscosity was similar at C and L (11.2, 10.2 cps) but somewhat higher at D (14.4 cps). Crude protein (6.25 N) ranged from 116 to 147 g/kg and tended to be higher at D. Starch, which ranged from 612 to 656 g/kg, was least at D (617 g/kg) and greatest at L (641 g/kg). 4. Crude protein, crude fibre and total non-starch polysaccharide (NSP) were negatively correlated with SW, the R2 being respectively 0.38 (P<0.05), 0.16 (NS) and 0.45 (P<0.05). TGW and starch concentration were positively correlated with SW (R2=0.70, 0.44, respectively). There was a weak (NS) negative relationship (R2=0.19) between in vitro viscosity and SW. For both TGW and in vitro viscosity, correlations improved when variety was taken into account (R2=0.95, 0.92, respectively). 5. There were no significant effects of variety on dry matter (DM) intake or live weight gain (LWG). Gain: food was significantly higher (P<0.05) for Consort than for the other three varieties and the metabolisable energy ratio (ME:GE) just failed to attain significance (P=0.062). Calculated wheat AME (MJ/kg DM) was significantly (P<0.05) higher for Consort than for the other three wheats. There was a good correlation (R2=0.49) for the total data set between gain:food and ME:GE. In vivo viscosity varied from 13.6 to 28.6 cps for individual treatments and was significantly affected by variety (P<0001). 6. Although there were no significant differences in DM intake or LWG due to site the values for L (SW 76) were 6 and 5% lower, respectively, than for D (SW 66). Gain:food was lower (P<0.05) for C (SW 69) than for D. ME:GE, wheat AME and ME:gain were not significantly different between sites. 7. There was a weak (R2=0.18) positive relationship between ME:GE and SW corresponding to a 2.5% increase in energy value for a 10 kg/hl increase in SW and no relationship between gain:food and SW. When variety was taken into the regression the slope was similar but R2 increased to 0.82. 8. ME:GE and wheat AME concentration were negatively correlated with wheat in vitro viscosity (R2=0.64, 0.55, respectively). 9. It was concluded that in vitro viscosity appears to provide a better basis than SW for prediction of the nutritive value of wheats of unknown variety. If the variety is known then SW could be used to predict energy value. However, the effect of quite a large change in SW (10 kg/hl) was relatively small.

Animal Feed↗
PubMed2001

Latency associated promoter transgene expression in the central nervous system after stereotaxic delivery of replication-defective HSV-1-based vectors.

The herpes simplex virus type 1 (HSV-1) latency associated promoter (LAP) has been shown to sustain long-term reporter gene expression within sensory neurones. Its activity within the CNS is, however, less well understood. In this study we characterise the activity of the LAP after stereotaxic delivery of recombinant HSV-1-based vectors to the brain. Two classes of vectors were utilised in these studies: (1) a replication-defective vector lacking the glycoprotein H and thymidine kinase genes, designated CS1, and (2) a virus mutant severely impaired for immediate-early (IE) gene expression which lacks functional VP16, ICP4 and ICP0 genes, designated in1388. Both vectors contain the LacZ gene under the control of the LAP. Following delivery of either vector to the striatum, beta-gal expression was detected within anatomically related CNS regions distal to the site of injection. At these sites the number of beta-gal-positive cells increased with time and remained stable up to 4 weeks p.i. beta-Gal expression could not be detected at the site of injection after delivery of CS1 but beta-gal expression within neurones located at this site was observed after delivery of in1388, indicating reduced toxicity of this severely disabled virus. Transgene expression decreased dramatically with both vectors at later time-points (>4 weeks after delivery), but PCR analysis demonstrated that viral genomes were stably maintained for up to 180 days following delivery, indicating that the loss of beta-gal-positive neurones was not likely to be due to a loss of vector-transduced cells. Moreover, after delivery of an equivalent virus to the rat striatum in situ hybridisation analysis showed a similar decrease in the number of neurones expressing the endogenous LATs with time. These data indicate that although the HSV-1 LAP can drive the expression of foreign genes in a variety of CNS neurones, in these cells there is a slow down-regulation of the viral promoter which eventually results in the loss of detectable transgene expression.

Animals↗
PubMed2001

Effect of wheat content, fat source and enzyme supplementation on diet metabolisability and broiler performance.

1. A 3x3x2 factorial experiment studied the interactions of fat source (tallow, soya, tallow:soya [2:1] blend), wheat level (700, 350, 0 g/kg) and enzyme inclusion (Avizyme 1300, absent, present) in diets for broilers fed ad libitum in individual cages from 7 to 35d. Bird performance, fat digestibility, viscosity of ileal contents and diet metabolisability (AME) were measured. 2. There were no significant effects of fat source on bird performance. However, there was a significant effect on fat digestibility, which was highest for soya and lowest for tallow. Diet AME content was also significantly affected by fat source and reflected differences in fat digestibility. 3. Dry matter (DM) intake, liveweight gain (LWG) and gain:food were all reduced at 700 g wheat/kg. Viscosity of ileal contents increased with increasing wheat inclusion. 4. There were no significant effects of enzyme on DM intake or LWG but gain:food was improved by 2%. Diet AME content was increased with enzyme addition, the effect being greatest (9%) with tallow at 700 g wheat/kg. 5. Viscosity of ileal contents was reduced and fat digestibility increased with enzyme addition and there were significant wheat enzyme interactions attributable to no differences with zero wheat but marked responses to enzyme at 700 g wheat/kg. 6. The results confirm important interactions between wheat content and fat composition in relation to fat digestibility, AME content and food efficiency.

Animals↗
PubMed2001

Activation of interferon response factor-3 in human cells infected with herpes simplex virus type 1 or human cytomegalovirus.

Activation of cellular interferon-stimulated genes (ISGs) after infection with herpes simplex virus type 1 (HSV-1) or human cytomegalovirus (HCMV) was investigated. The level of ISG54-specific RNA in human fetal lung (HFL) or human foreskin (BJ) fibroblasts increased substantially after infection with either virus in the presence of cycloheximide. HSV-1 particles lacking glycoprotein D or glycoprotein H failed to induce ISG54-specific RNA synthesis, demonstrating that entry of virus particles rather than binding of virions to the cell surface was required for the effect. A DNA-binding complex that recognized an interferon-responsive sequence motif was induced upon infection with HSV-1 or HCMV in the presence of cycloheximide, and the complex was shown to contain the cell proteins interferon response factor 3 (IRF-3) and CREB-binding protein. IRF-3 was modified after infection with HSV-1 or HCMV to a form of lower electrophoretic mobility, consistent with phosphorylation. De novo transcription of viral or cellular genes was not required for the activation of IRF-3, since the effect was not sensitive to inhibition by actinomycin D. Infection of HFL fibroblasts with HSV-1 under conditions in which viral replication proceeded normally resulted in severely reduced levels of the IRF-3-containing complex, defining the activation of IRF-3 as a target for viral interference with ISG induction. In BJ fibroblasts, however, significant activation of IRF-3 was detected even when the viral gene expression program progressed to later stages, demonstrating that the degree of inhibition of the response was dependent on host cell type. As a consequence of IRF-3 activation, endogenous interferon was released from BJ cells and was capable of triggering the appropriate signal transduction pathway in both infected and uninfected cells. Activation of ISG54-specific RNA synthesis was not detected after infection of human U-373MG glioblastoma cells, showing that the induction of the response by infection is cell type dependent.

Apoptosis Regulatory Proteins↗
PubMed2001

Herpes simplex virus type 1 blocks the apoptotic host cell defense mechanisms that target Bcl-2 and manipulates activation of p38 mitogen-activated protein kinase to improve viral replication.

Wild-type (wt) herpes simplex virus type 1 (HSV-1) suppresses cell death. We investigated the apoptotic pathways triggered during infection with mutant viruses tsk and 27lacZ (which lack functional ICP4 and ICP27 viral proteins, respectively) and examined the mechanisms used by wt HSV-1 to protect against programmed cell death induced by the DNA-damaging compound cisplatin. In our studies, we used BHK and HeLa cells, with similar results. We suggest that a decrease in the levels of Bcl-2 protein is a key event during apoptosis induced by the mutant viruses and that Bcl-2 levels are targeted by (i) a decrease of bcl-2 RNA, (ii) caspase-related proteolysis, and (iii) p38 mitogen-activated protein kinase (p38MAPK)-dependent destabilization of Bcl-2 protein. We show that wt HSV-1, but not the mutant viruses, maintains bcl-2 RNA and protein levels during infection and protects from the cisplatin-induced decrease in bcl-2 RNA; our data suggest that both ICP27 and ICP4 are required for this function. Additionally, wt HSV-1 evades but does not actively block activation of caspases. Although wt HSV-1 induces p38MAPK activation during infection, it prevents p38MAPK-dependent destabilization of Bcl-2 and exploits p38MAPK stimulation to enhance transcription of specific viral gene promoters to increase viral yields.

Animals↗
PubMed2000

Subcellular post-transcriptional targeting: delivery of an intracellular protein to the extracellular leaflet of the plasma membrane using a glycosyl-phosphatidylinositol (GPI) membrane anchor in neurons and polarised epithelial cells.

The effectiveness of viral vector-mediated gene transfer depends on the expression of therapeutic transgenes in the correct target cell types. So far, however, little attention has been given to targeted subcellular distribution of expressed transgenes. Targeting individual transgenes to particular subcellular compartments will provide various advantages in increasing the safety, efficacy, and specificity of viral vector-mediated gene delivery. Viruses normally hijack the cellular protein synthesis machinery for their own advantages. It is thus unknown whether cells infected with viral vectors will be able to target proteins to the correct subcellular organelles, or whether the subcellular targeting machinery would be selectively disrupted by viral infection. In this article we explored whether a herpes simplex virus type 1-derived vector could be used to deliver a transgene engineered to be targeted to the extracellular membrane of target cells. To do so we constructed a temperature-sensitive mutant HSV-1 vector, tsK-TT21 expressing a recombinant marker protein, tissue inhibitor of metalloproteinases (TIMP), linked to sequence encoding a signal for the addition of a glycosyl-phosphatidylinositol (GPI)-anchor within the endoplasmic reticulum. Our results demonstrate that HSV1-derived viral vectors can be used to target transgenes as GPI anchored proteins to the outside leaflet of plasma membranes, without disrupting the targeting machinery of host epithelial cells or neurons. This approach could then be used to target specific proteins to the cell membrane to modify cell-cell interactions, the function of specific plasma membrane proteins, or their interactions with other membrane proteins, and also to target a prodrug converting enzyme to the plasma membrane of target cells, therefore enhancing its cell killing effects.

Biotinylation↗
PubMed2000

Effect of diet form and enzyme supplementation on growth, efficiency and energy utilisation of wheat-based diets for broilers.

1. The effect of diet form (mash, cold-pelleted, steam-conditioned/pelleted, wet mash, whole wheat with balancer pellet, restricted pellet) and enzyme inclusion (Avizyme 1300, absent, present) was studied in 2 trials using individually caged, male broilers from 14 to 42 d. Bird performance, viscosity of ileal contents and diet metabolisability (AME) were measured. 2. The performance of mash-fed birds was significantly poorer than for the other treatments in relation to dry matter intake, liveweight gain and gain:food. This was not due to reduced diet AME content. 3. There was no significant effect of heat treatment on any of the variables measured, although viscosity of ileal contents was increased by 30% as compared to the cold-pelleted diet. 4. Gain:food was improved with wet-mash feeding in comparison to the dry mash treatment but it was concluded that this was not due to any intrinsic improvement in diet quality, but rather to voluntary food restriction on introduction of the wet food. 5. Whole wheat feeding improved gain:food and diet AME content by 3% as compared to the complete diets and caused approximately a 50% increase in gizzard weight as compared with the pelleted diets. 6. Food enzyme inclusion did not improve performance although a significant improvement in diet AME content was observed with enzyme inclusion in trial 1.

Amino Acids↗
PubMed2000

Long-term transgene expression in mice infected with a herpes simplex virus type 1 mutant severely impaired for immediate-early gene expression.

The role of viral immediate-early (IE) gene expression in herpes simplex virus type 1 (HSV-1) latency was investigated. The HSV-1 multiple mutant in1312, defective for the expression of the virion transactivator VP16 and the IE proteins ICP0 and ICP4, was used as the parent for these studies. The coding sequences of the Escherichia coli lacZ gene, preceded by the encephalomyocarditis virus internal ribosome entry site, were inserted into the region of in1312 that encodes the latency-associated transcripts (LATs) such that transcription of the transgene was controlled by the LAT promoter. This insert has previously been shown to direct long-term latent-phase expression of beta-galactosidase in a wild-type HSV-1 genome (R. H. Lachmann and S. Efstathiou, J. Virol. 71, 3197-3207, 1997). The resulting recombinant, in1388, was apathogenic after inoculation into mice via the footpad and did not detectably replicate in dorsal root ganglia (DRG) or footpads. Mutant in1388 established latency in DRG, and beta-galactosidase was expressed in increasing numbers of neurons over the first 25 days of infection. During latency, more than 1% of neurons in ganglia that innervate the footpad expressed beta-galactosidase, with the number of positive cells remaining constant for at least 5 months. Rescue of the VP16, ICP0, or ICP4 mutations of in1388 did not affect the number of beta-galactosidase-expressing neurons detected during latency. The results demonstrate that HSV-1 mutants severely impaired for IE gene expression are capable of establishing latency and efficiently expressing a foreign gene product under control of the LAT promoter.

Animals↗
PubMed1999

A non-cytotoxic herpes simplex virus vector which expresses Cre recombinase directs efficient site specific recombination.

The coding sequences for the bacteriophage P1 recombinase Cre were cloned into the genome of a herpes simplex virus type 1 (HSV-1) mutant which is severely impaired for the synthesis of immediate early (IE) proteins. The resulting recombinant, virus in1372, expressed functional Cre which mediated the excision in trans of loxP-flanked sequences located in the HSV-1 genome, both in tissue culture cells and in vivo in mouse sensory neurons. Infection with in1372 also resulted in recombination, at high efficiency, between loxP sequences in the cellular genome without causing detectable cytotoxicity. Mutant in1372 is a versatile vector for the delivery of Cre in tissue culture and in vivo.

Animals↗
PubMed1999

Visualization and enumeration of marine planktonic archaea and bacteria by using polyribonucleotide probes and fluorescent in situ hybridization.

Fluorescent in situ hybridization (FISH) using rRNA-specific oligonucleotide probes has emerged as a popular technique for identifying individual microbial cells. In natural samples, however, the signal derived from fluor-labeled oligonucleotide probes often is undetectable above background fluorescence in many cells. To circumvent this difficulty, we applied fluorochrome-labeled polyribonucleotide probes to identify and enumerate marine planktonic archaea and bacteria. The approach greatly enhanced the sensitivity and applicability of FISH with seawater samples, allowing confident identification and enumeration of planktonic cells to ocean depths of 3,400 m. Quantitative whole-cell hybridization experiments using these probes accounted for 90 to 100% of the total 4',6-diamidino-2-phenylindole (DAPI)-stained cells in most samples. As predicted in a previous study (R. Massana, A. E. Murray, C. M. Preston, and E. F. DeLong, Appl. Environ. Microbiol. 63:50-56, 1997), group I and II marine archaea predominate in different zones in the water column, with maximal cell densities of 10(5)/ml. The high cell densities of archaea, extending from surface waters to abyssal depths, suggest that they represent a large and significant fraction of the total picoplankton biomass in coastal ocean waters. The data also show that the vast majority of planktonic prokaryotes contain significant numbers of ribosomes, rendering them easily detectable with polyribonucleotide probes. These results imply that the majority of planktonic cells visualized by DAPI do not represent lysed cells or "ghosts," as was suggested in a previous report.

Animals↗
PubMed1999

Activation of herpesvirus gene expression by the human cytomegalovirus protein pp71.

The activation of gene expression by the human cytomegalovirus (HCMV) particle was investigated. The HCMV major immediate-early (IE) promoter was cloned upstream of the Escherichia coli lacZ coding sequences, and the resulting cassette was introduced into the genome of a herpes simplex virus type 1 (HSV-1) mutant lacking functional VP16. Upon infection with the HSV-1 recombinant in the presence of cycloheximide, to block de novo protein synthesis, expression of lacZ-specific transcripts was increased by fivefold when HCMV was included in the inoculum. Accumulation of HSV-1 IE RNAs was also stimulated by coinfection with HCMV, as was expression of the adenovirus 5 VAI transcript when the VAI gene was cloned into the HSV-1 genome. Coinfection with HCMV did not alter mRNA stability or uncoating of the HSV-1 genome. The coding sequences for the HCMV phosphoprotein pp71, controlled by the HCMV IE promoter, were cloned into an HSV-1 recombinant impaired for the production of the three major transactivators (VP16, ICP0, and ICP4) to yield a recombinant (in1324) which expressed pp71 but did not cause significant cytotoxicity. Infection with in1324 resulted in stimulation of HCMV IE, HSV-1 IE, and VAI expression, demonstrating that pp71 is responsible for the effects we observed when using the entire HCMV particle. Therefore, HCMV pp71 exhibits novel properties in its ability to stimulate gene expression from a range of promoters present in a herpesvirus genome.

Cell Line↗